We propose to define the structural aspects of the human RBC membrane responsible for Na ion and K ion permeability. Our hypothesis is that the low molecular weight membrane polypeptides designated bands 7.1, 7.2, 7.3 and 8 form permeability channels which are aberrant in clinical disorders of RBC cation and water balance. Others have demonstrated that bands 7 and 8 associate in low ionic strength RBC extracts to form a 747,000 dalton protein with a "hollow cylinder" appearance on electron microscopy. We have documented an abnormal band 7 in two patients with primary defects of RBC permeability. We now plan to examine pathologic RBC with primary permeability defects as well as RBC with secondary defects due to hemoglobinopathies or metabolic depletion. The "hollow cylinder" protein (HCP) and band 7 and 8 in such pathologic RBC will be studied by transmission electron microscopy and by two-dimensional electrophoresis. The partition of the HCP between the cytosol and the membrane, and the membrane organization of the HCP will be evaluated. Selective extractions, cross-linking studies with sulfhydral and amino reacting reagents, and surface labeling by iodination and hemoglobin cross-linking will be carried out. The topographic localization of the HCP will be examined by transmission electron microscopy utilizing ferritin-conjugated antibody to the HCP and freeze-etched membranes from pathologic RBC. The relationship of the HCP to the sodium and potassium permeability channel will be assessed by radiolabeled imidoester chemical probes, and by the effect of specific antibody on the permeability of right-side out and inside-out membranes. Such studies should define the structural basis for Na ion and K ion permeability in normal RBC membranes and indicate how inherited abnormalities of membrane proteins may adversely affect permeability.